Prolonged treatment of chick embryos in vivo with the muscarinic agonists oxotremorine or carbachol leads to dose- and time-dependent decreases in the number of brain muscarinic acetylcholine receptors (mAChR) as measured by the specific binding of the potent muscarinic ligand L-[3H]quinuclidinyl benzilate to brain membranes. Maximal doses of agonists reduced the number of mAChR as much as 44%. Maximal loss of mAChR occurs 4 hr after treatment, but can be prevented or totally reversed within 24 hr by blockade of agonist-receptor interactions with muscarinic antagonists. After sustained in vivo oxotremorine treatment, brain mAChR show a decreased apparent affinity for agonists owing to a decrease in the affinities of both the high- and low-affinity agonist binding sites.